Effects of Cross-linker Chemistry on Bioelectrocatalytic Reactions in a Redox Cross-linked Network of Glucose Dehydrogenase and Thionine.

Enzyme-mediator bioconjugation is emerging as a building block for designing electrode platforms for the construction of biosensors and biofuel cells. Here, we report a one-pot bioconjugation technique for flavin adenine dinucleotide-dependent glucose dehydrogenase (FAD-GDH) and thionine (TH) using a series of cross-linkers, including epoxy, N-hydroxysuccinimide (NHS), and aldehydes. In this technique, FAD-GDH and thionine are conjugated through an amine cross-linking reaction to generate a redox network, which has been successfully employed for the oxidation of glucose. The bioconjugation chemistry of cross-linkers with the amino groups on FAD-GDH and thionine plays a vital role in generating distinct network structures. The epoxy-type cross-linker reacts with the primary and secondary amines of thionine at room temperature, thereby producing an FAD-GDH-TH-FAD-GDH hyperbranched bioconjugate network, the aldehyde undergoes a rapid cross-linking reaction to produce a network of FAD-GDH-FAD-GDH, while the NHS-based cross-linker can react with the primary amines of both FAD-GDH and thionine, forming an FAD-GDH-cross-linker-TH polymeric network. This reaction has the potential to enable the conjugation of a redox mediator with a FAD-GDH network, which is particularly essential when designing an enzyme electrode platform. The data demonstrated that the polymeric cross-linked network based on the NHS cross-linker exhibited a considerable increase in electron transport while producing a catalytic current of 830 µA cm-2. The cross-linker spacer arm length also affects the overall electrochemical function of the network and its performance; an adequate spacer length containing a cross-linker is required, resulting in a faster electron transfer. Finally, a leaching test confirmed that the stability of the enzyme electrode was improved when the electrode was tested using the redox probe. This study elucidates the relationship between cross-linking chemistry and redox network structure and enhances the high performance of enzyme electrode platforms for the oxidation of glucose.

Synthetic Nicotinamide Cofactors as Alternatives to NADPH in Imine Reductase-Catalyzed Reactions.

This study demonstrates the effectiveness of synthetic nicotinamide cofactors as cost-effective alternatives to NADPH in imine reductase (IRED) catalysis. The synthetic cofactors maintained catalytic activity and stereoselectivity, achieving high conversion rates. Molecular docking studies revealed key structural interactions influencing performance. Combining a glucose dehydrogenase (GDH) recycling system further enhanced the stability and efficiency. These findings highlight the potential of synthetic cofactors to reduce costs and improve the feasibility of IRED-catalyzed processes for industrial applications.

Bias-free glucose/O2 bio-photoelectrochemical system for multi-energy conversion and phenolic pollutant degradation.

Developing a multi-functional green energy device that propels sustainable energy development and concurrently purifies environmental pollutants offers an irresistibly compelling vision for a cleaner future. Herein, we reported a bias-free glucose/O2 bio-photoelectrochemical system (BPECS) for both energy conversion and phenolic pollutants degradation. Coupling a glucose dehydrogenase (GDH) modified self-assembled meso-tetrakis (4-carboxyphenyl)-porphyrin (SA-TCPP)-sensitized TiO2 biophotoanode for glucose oxidation and nitrogen/oxygen doped cobalt single-atom catalyst (CoNOC) cathode for two-electron oxygen reduction, both solar and biochemical energies were converted into electric power in BPECS with a maximum power density of 296.98 µW cm-2 (0.49 V). Working in synergy with horseradish peroxidase (HRP) biocatalysis, the cathode-generated H2O2, a by-product, is effectively redeployed for degrading phenol, attaining an impressive degradation efficiency of approximately 100% within 60 min. Additionally, aiming to scale up this ingenious BPECS approach, peroxidase-mimicking Co3O4 nanozyme were engineered as a substitute for natural HRP. Remarkably, these nanozyme demonstrated a comparable degradation efficiency, achieving the same result in 90 min. In this work, our results demonstrate that this bias-free glucose/O2 BPECS model marks a significant step forward in integrating renewable energy harvesting with environmental remediation, but also opens new avenues for the versatile application of nanozymes.

Shifting redox reaction equilibria on demand using an orthogonal redox cofactor.

Nature's two redox cofactors, nicotinamide adenine dinucleotide (NAD+) and nicotinamide adenine dinucleotide phosphate (NADP+), are held at different reduction potentials, driving catabolism and anabolism in opposite directions. In biomanufacturing, there is a need to flexibly control redox reaction direction decoupled from catabolism and anabolism. We established nicotinamide mononucleotide (NMN+) as a noncanonical cofactor orthogonal to NAD(P)+. Here we present the development of Nox Ortho, a reduced NMN+ (NMNH)-specific oxidase, that completes the toolkit to modulate NMNH:NMN+ ratio together with an NMN+-specific glucose dehydrogenase (GDH Ortho). The design principle discovered from Nox Ortho engineering and modeling is facilely translated onto six different enzymes to create NMN(H)-orthogonal biocatalysts with a consistent ~103-106-fold cofactor specificity switch from NAD(P)+ to NMN+. We assemble these enzymes to produce stereo-pure 2,3-butanediol in cell-free systems and in Escherichia coli, enabled by NMN(H)'s distinct redox ratio firmly set by its designated driving forces, decoupled from both NAD(H) and NADP(H).

Sources and control of impurity during one-pot enzymatic production of dehydroepiandrosterone.

Dehydroepiandrosterone (DHEA) has a promising market due to its capacity to regulate human hormone levels as well as preventing and treating various diseases. We have established a chemical esterification coupled biocatalytic-based scheme by lipase-catalyzed 4-androstene-3,17-dione (4-AD) hydrolysis to obtain the intermediate product 5-androstene-3,17-dione (5-AD), which was then asymmetrically reduced by a ketoreductase from Sphingomonas wittichii (SwiKR). Co-enzyme required for KR is regenerated by a glucose dehydrogenase (GDH) from Bacillus subtilis. This scheme is more environmentally friendly and more efficient than the current DHEA synthesis pathway. However, a significant amount of 4-AD as by-product was detected during the catalytic process. Focused on the control of by-products, we investigated the source of 4-AD and identified that it is mainly derived from the isomerization activity of SwiKR and GDH. Increasing the proportion of glucose in the catalytic system as well as optimizing the catalytic conditions drastically reduced 4-AD from 24.7 to 6.5% of total substrate amount, and the final yield of DHEA achieved 40.1 g/L. Furthermore, this is the first time that both SwiKR and GDH have been proved to be promiscuous enzymes with dehydrogenase and ketosteroid isomerase (KSI) activities, expanding knowledge of the substrate diversity of the short-chain dehydrogenase family enzymes. KEY POINTS: • A strategy of coupling lipase, ketoreductase, and glucose dehydrogenase in producing DHEA from 4-AD • Both SwiKR and GDH are identified with ketosteroid isomerase activity. • Development of catalytic strategy to control by-product and achieve highly selective DHEA production.

A Multienzyme Logic H+ and Na+ Biotransducer.

Sodium ions and protons regulate various fundamental processes at the cell and tissue levels across all biological kingdoms. It is therefore pivotal for bioelectronic devices, such as biosensors and biotransducers, to control the transport of these ions through biological membranes. Our study explores the regulation of proton and sodium concentrations by integrating an Na+-type ATP synthase, a glucose dehydrogenase (GDH), and a urease into a multienzyme logic system. This system is designed to operate using various chemical control input signals, while the output current corresponds to the local change in proton or sodium concentrations. Therein, a H+ and Na+ biotransducer was integrated to fulfill the roles of signal transducers for the monitoring and simultaneous control of Na+ and H+ levels, respectively. To increase the proton concentration at the output, we utilized GDH driven by the inputs of glucose and nicotinamide adenine dinucleotide (NAD+), while recorded the signal change from the biotransducer, together acting as an AND enzyme logic gate. On the contrary, we introduced urease enzyme which hydrolyzed urea to control the decrease in proton concentration, serving as a NOT gate and reset. By integrating these two enzyme logic gates we formed a simple multienzyme logic system for the control of proton concentrations. Furthermore, we also demonstrate a more complex, Na+-type ATP synthase-urease multienzyme logic system, controlled by the two different inputs of ADP and urea. By monitoring the voltage of the peak current as the output signal, this logic system acts as an AND enzyme logic gate. This study explores how multienzyme logic systems can modulate biologically important ion concentrations, opening the door toward advanced biological on-demand control of a variety of bioelectronic enzyme-based devices, such as biosensors and biotransducers.

Nanoarchitectonics of Vesicle Microreactors for Oscillating ATP Synthesis and Hydrolysis.

We construct a compartmentalized nanoarchitecture to regulate bioenergy level. Glucose dehydrogenase, urease and nicotinamide adenine dinucleotide are encapsulated inside through liquid-liquid phase separation. ATPase and glucose transporter embedded in hybrid liposomes are attached at the surface. Glucose is transported and converted to gluconic acid catalyzed by glucose dehydrogenase, resulting in an outward proton gradient to drive ATPase for ATP synthesis. In parallel, urease catalyzes hydrolysis of urea to generate ammonia, which leads to an inward proton gradient to drive ATPase for ATP hydrolysis. These processes lead to a change of the direction of proton gradient, thus achieving artificial ATP oscillation. Importantly, the frequency and the amplitude of the oscillation can be programmed. The work explores nanoarchitectonics integrating multiple components to realize artificial and precise oscillation of bioenergy level.

Harnessing Redox Polymer Dynamics for Enhanced Glucose-Oxygen Coupling in Dual Biosensing and Therapeutic Applications.

The burgeoning field of continuous glucose monitoring (CGM) for diabetes management faces significant challenges, particularly in achieving precise and stable biosensor performance under changing environmental conditions such as varying glucose concentrations and O2 levels. To address this, we present a novel biosensor based on the electroless coupling of glucose oxidation catalyzed by flavin-dependent glucose dehydrogenase (FAD-GDH) and O2 reduction catalyzed by bilirubin oxidase (BOD) via a redox polymer, dimethylferrocene-modified linear poly(ethylenimine), FcMe2-LPEI. Initial cyclic voltammetry tests confirm the colocalization of both enzymatic reactions within the potential range of the polymer, indicating an effective electron shuttle mechanism. As a result, we created a hybrid biosensor that operates at open-circuit potential (OCP). It can detect glucose concentrations of up to 100 mM under various O2 conditions, including ambient air. This resulted from optimizing the enzyme ratio to 120 ± 10 mUBOD·UFAD-GDH-1·atmO2-1. This biosensor is highly sensitive, a crucial feature for CGM applications. This distinguishes it from FAD-GDH traditional biosensors, which require a potential to be applied to measure glucose concentrations up to 30 mM. In addition, this biosensor demonstrates the ability to function as a noninvasive, external device that can adapt to changing glucose levels, paving the way for its use in diabetes care and, potentially, personalized healthcare devices. Furthermore, by leveraging the altered metabolic pathways in tumor cells, this system architecture opened up new avenues for targeted glucose scavenging and O2 reduction in cancer therapy.

Coupling protein scaffold and biosilicification: A sustainable and recyclable approach for d-mannitol production via one-step purification and immobilization of multienzymes.

Enzymatic synthesis of biochemicals in vitro is vital in synthetic biology for its efficiency, minimal by-products, and easy product separation. However, challenges like enzyme preparation, stability, and reusability persist. Here, we introduced a protein scaffold and biosilicification coupled system, providing a singular process for the purification and immobilization of multiple enzymes. Using d-mannitol as a model, we initially constructed a self-assembling EE/KK protein scaffold for the co-immobilization of glucose dehydrogenase and mannitol dehydrogenase. Under an enzyme-to-scaffold ratio of 1:8, a d-mannitol yield of 0.692 mol/mol was achieved within 4 h, 2.16-fold higher than the free enzymes. The immobilized enzymes retained 70.9 % of the initial joint activity while the free ones diminished nearly to inactivity after 8 h. Furthermore, we incorporated the biosilicification peptide CotB into the EE/KK scaffold, inducing silica deposition, which enabled the one-step purification and immobilization process assisted by Spy/Snoop protein-peptide pairs. The coupled system demonstrated a comparable d-mannitol yield to that of EE/KK scaffold and 1.34-fold higher remaining activities after 36 h. Following 6 cycles of reaction, the immobilized system retained the capability to synthesize 56.4 % of the initial d-mannitol titer. The self-assembly co-immobilization platform offers an effective approach for enzymatic synthesis of d-mannitol and other biochemicals.

Co-Immobilization of ADH and GDH on Metal-Organic-Framework: An Effective Biocatalyst for Asymmetric Reduction of Ketones.

Chiral alcohols are not only important building blocks of various bioactive natural compounds and pharmaceuticals, but can serve as synthetic precursors for other valuable organic chemicals, thus the synthesis of these products is of great importance. Bio-catalysis represents one effective way to obtain these molecules, however, the weak stability and high cost of enzymes often hinder its broad application. In this work, we designed a biological nanoreactor by embedding alcohol dehydrogenase (ADH) and glucose dehydrogenase (GDH) in metal-organic-framework ZIF-8. The biocatalyst ADH&GDH@ZIF-8 could be applied to the asymmetric reduction of a series of ketones to give chiral alcohols in high yields (up to 99 %) and with excellent enantioselectivities (>99 %). In addition, the heterogeneous biocatalyst could be recycled and reused at least four times with slight activity decline. Moreover, E. coli containing ADH and GDH was immobilized by ZIF-8 to form biocatalyst E. coli@ZIF-8, which also exhibits good catalytic behaviours. Finally, the chiral alcohols are further converted to marketed drugs (R)-Fendiline, (S)-Rivastigmine and NPS R-568 respectively.

Does Acinetobacter calcoaceticus glucose dehydrogenase produce self-damaging H2O2?

The soluble glucose dehydrogenase (sGDH) from Acinetobacter calcoaceticus has been widely studied and is used, in biosensors, to detect the presence of glucose, taking advantage of its high turnover and insensitivity to molecular oxygen. This approach, however, presents two drawbacks: the enzyme has broad substrate specificity (leading to imprecise blood glucose measurements) and shows instability over time (inferior to other oxidizing glucose enzymes). We report the characterization of two sGDH mutants: the single mutant Y343F and the double mutant D143E/Y343F. The mutants present enzyme selectivity and specificity of 1.2 (Y343F) and 5.7 (D143E/Y343F) times higher for glucose compared with that of the wild-type. Crystallographic experiments, designed to characterize these mutants, surprisingly revealed that the prosthetic group PQQ (pyrroloquinoline quinone), essential for the enzymatic activity, is in a cleaved form for both wild-type and mutant structures. We provide evidence suggesting that the sGDH produces H2O2, the level of production depending on the mutation. In addition, spectroscopic experiments allowed us to follow the self-degradation of the prosthetic group and the disappearance of sGDH's glucose oxidation activity. These studies suggest that the enzyme is sensitive to its self-production of H2O2. We show that the premature aging of sGDH can be slowed down by adding catalase to consume the H2O2 produced, allowing the design of a more stable biosensor over time. Our research opens questions about the mechanism of H2O2 production and the physiological role of this activity by sGDH.

Catalytic Synthesis of (S)-CHBE by Directional Coupling and Immobilization of Carbonyl Reductase and Glucose Dehydrogenase.

Ethyl (S)-4-chloro-3-hydroxybutyrate ((S)-CHBE) is an important chiral intermediate in the synthesis of the cholesterol-lowering drug atorvastatin. Studying the use of SpyTag/SpyCatcher and SnoopTag/SnoopCatcher systems for the asymmetric reduction reaction and directed coupling coenzyme regeneration is practical for efficiently synthesizing (S)-CHBE. In this study, Spy and Snoop systems were used to construct a double-enzyme directed fixation system of carbonyl reductase (BsCR) and glucose dehydrogenase (BsGDH) for converting 4-chloroacetoacetate (COBE) to (S)-CHBE and achieving coenzyme regeneration. We discussed the enzymatic properties of the immobilized enzyme and the optimal catalytic conditions and reusability of the double-enzyme immobilization system. Compared to the free enzyme, the immobilized enzyme showed an improved optimal pH and temperature, maintaining higher relative activity across a wider range. The double-enzyme immobilization system was applied to catalyze the asymmetric reduction reaction of COBE, and the yield of (S)-CHBE reached 60.1% at 30 °C and pH 8.0. In addition, the double-enzyme immobilization system possessed better operational stability than the free enzyme, and maintained about 50% of the initial yield after six cycles. In summary, we show a simple and effective strategy for self-assembling SpyCatcher/SnoopCatcher and SpyTag/SnoopTag fusion proteins, which inspires building more cascade systems at the interface. It provides a new method for facilitating the rapid construction of in vitro immobilized multi-enzyme complexes from crude cell lysate.

Development of Direct Electron Transfer-Type Extended Gate Field Effect Transistor Enzymatic Sensors for Metabolite Detection.

In this work, direct electron transfer (DET)-type extended gate field effect transistor (EGFET) enzymatic sensors were developed by employing DET-type or quasi-DET-type enzymes to detect glucose or lactate in both 100 mM potassium phosphate buffer and artificial sweat. The system employed either a DET-type glucose dehydrogenase or a quasi-DET-type lactate oxidase, the latter of which was a mutant enzyme with suppressed oxidase activity and modified with amine-reactive phenazine ethosulfate. These enzymes were immobilized on the extended gate electrodes. Changes in the measured transistor drain current (ID) resulting from changes to the working electrode junction potential (φ) were observed as glucose and lactate concentrations were varied. Calibration curves were generated for both absolute measured ID and ΔID (normalized to a blank solution containing no substrate) to account for variations in enzyme immobilization and conjugation to the mediator and variations in reference electrode potential. This work resulted in a limit of detection of 53.9 µM (based on ID) for glucose and 2.12 mM (based on ID) for lactate, respectively. The DET-type and Quasi-DET-type EGFET enzymatic sensor was then modeled using the case of the lactate sensor as an equivalent circuit to validate the principle of sensor operation being driven through OCP changes caused by the substrate-enzyme interaction. The model showed slight deviation from collected empirical data with 7.3% error for the slope and 8.6% error for the y-intercept.

Ketogluconate production by Gluconobacter strains: enzymes and biotechnological applications.

Gluconobacter strains perform incomplete oxidation of various sugars and alcohols, employing regio- and stereoselective membrane-bound dehydrogenases oriented toward the periplasmic space. This oxidative fermentation process is utilized industrially. The ketogluconate production pathway, characteristic of these strains, begins with the conversion of d-glucose to d-gluconate, which then diverges and splits into 2 pathways producing 5-keto-d-gluconate and 2-keto-d-gluconate and subsequently 2,5-diketo-d-gluconate. These transformations are facilitated by membrane-bound d-glucose dehydrogenase, glycerol dehydrogenase, d-gluconate dehydrogenase, and 2-keto-d-gluconate dehydrogenase. The variance in end products across Gluconobacter strains stems from the diversity of enzymes and their activities. This review synthesizes biochemical and genetic knowledge with biotechnological applications, highlighting recent advances in metabolic engineering and the development of an efficient production process focusing on enzymes relevant to the ketogluconate production pathway in Gluconobacter strains.

Enzymatic Galvanic Redox Potentiometry for In Vivo Biosensing.

Redox potentiometry has emerged as a new platform for in vivo sensing, with improved neuronal compatibility and strong tolerance against sensitivity variation caused by protein fouling. Although enzymes show great possibilities in the fabrication of selective redox potentiometry, the fabrication of an enzyme electrode to output open-circuit voltage (EOC) with fast response remains challenging. Herein, we report a concept of novel enzymatic galvanic redox potentiometry (GRP) with improved time response coupling the merits of the high selectivity of enzyme electrodes with the excellent biocompatibility and reliability of GRP sensors. With a glucose biosensor as an illustration, we use flavin adenine dinucleotide-dependent glucose dehydrogenase as the recognition element and carbon black as the potential relay station to improve the response time. We find that the enzymatic GRP biosensor rapidly responds to glucose with a good linear relationship between EOC and the logarithm of glucose concentration within a range from 100 µM to 2.65 mM. The GRP biosensor shows high selectivity over O2 and coexisting neurochemicals, good reversibility, and sensitivity and can in vivo monitor glucose dynamics in rat brain. We believe that this study will pave a new platform for the in vivo potentiometric biosensing of chemical events with high reliability.

Sustainable asymmetric synthesis of diltiazem precursor enabled by recombinant Escherichia coli whole cells co-expressing an engineered ketoreductase and glucose dehydrogenase.

As a key synthetic intermediate of the cardiovascular drug diltiazem, methyl (2R,3S)-3-(4-methoxyphenyl) glycidate ((2R,3S)-MPGM) (1) is accessible via the ring closure of chlorohydrin (3S)-methyl 2-chloro-3-hydroxy-3-(4-methoxyphenyl)propanoate ((3S)-2). We report the efficient reduction of methyl 2-chloro-3-(4-methoxyphenyl)-3-oxo-propanoate (3) to (3S)-2 using an engineered enzyme SSCRM2 possessing 4.5-fold improved specific activity, which was obtained through the structure-guided site-saturation mutagenesis of the ketoreductase SSCR by reliving steric hindrance and undesired interactions. With the combined use of the co-expression fine-tuning strategy, a recombinant E. coli (pET28a-RBS-SSCRM2 /pACYCDuet-GDH), co-expressing SSCRM2 and glucose dehydrogenase, was constructed and optimized for protein expression. After optimizing the reaction conditions, whole-cell-catalyzed complete reduction of industrially relevant 300 g L-1 of 3 was realized, affording (3S)-2 with 99% ee and a space-time yield of 519.1 g∙L-1 ∙d-1 , representing the highest record for the biocatalytic synthesis of (3S)-2 reported to date. The E-factor of this biocatalytic synthesis was 24.5 (including water). Chiral alcohol (3S)-2 generated in this atom-economic synthesis was transformed to (2R,3S)-MPGM in 95% yield with 99% ee.

Characterization and Application of a Novel Glucose Dehydrogenase with Excellent Organic Solvent Tolerance for Cofactor Regeneration in Carbonyl Reduction.

An efficient cofactor regeneration system has been developed to provide a hydride source for the preparation of optically pure alcohols by carbonyl reductase-catalyzed asymmetric reduction. This system employed a novel glucose dehydrogenase (BcGDH90) from Bacillus cereus HBL-AI. The gene encoding BcGDH90 was found through the genome-wide functional annotation. Homology-built model study revealed that BcGDH90 was a homo-tetramer, and each subunit was composed of ßD-αE-αF-αG-ßG motif, which was responsible for substrate binding and tetramer formation. The gene of BcGDH90 was cloned and expressed in Escherichia coli. The recombinant BcGDH90 exhibited maximum activity of 45.3 U/mg at pH 9.0 and 40 °C. BcGDH90 showed high stability in a wide pH range of 4.0-10.0 and was stable after the incubation at 55 °C for 5 h. BcGDH90 was not a metal ion-dependent enzyme, but Zn2+ could seriously inhibit its activity. BcGDH90 displayed excellent tolerance to 90% of acetone, methanol, ethanol, n-propanol, and isopropanol. Furthermore, BcGDH90 was applied to regenerate NADPH for the asymmetric biosynthesis of (S)-(+)-1-phenyl-1,2-ethanediol ((S)-PED) from hydroxyacetophenone (2-HAP) with high concentration, which increased the final efficiency by 59.4%. These results suggest that BcGDH90 is potentially useful for coenzyme regeneration in the biological reduction.

Development of a Versatile Method to Construct Direct Electron Transfer-Type Enzyme Complexes Employing SpyCatcher/SpyTag System.

The electrochemical enzyme sensors based on direct electron transfer (DET)-type oxidoreductase-based enzymes are ideal for continuous and in vivo monitoring. However, the number and types of DET-type oxidoreductases are limited. The aim of this research is the development of a versatile method to create a DET-type oxidoreductase complex based on the SpyCatcher/SpyTag technique by preparing SpyCatcher-fused heme c and SpyTag-fused non-DET-type oxidoreductases, and by the in vitro formation of DET-type oxidoreductase complexes. A heme c containing an electron transfer protein derived from Rhizobium radiobacter (CYTc) was selected to prepare SpyCatcher-fused heme c. Three non-DET-type oxidoreductases were selected as candidates for the SpyTag-fused enzyme: fungi-derived flavin adenine dinucleotide (FAD)-dependent glucose dehydrogenase (GDH), an engineered FAD-dependent d-amino acid oxidase (DAAOx), and an engineered FMN-dependent l-lactate oxidase (LOx). CYTc-SpyCatcher (CYTc-SC) and SpyTag-Enzymes (ST-GDH, ST-DAAOx, ST-LOx) were prepared as soluble molecules while maintaining their redox properties and catalytic activities, respectively. CYTc-SC/ST-Enzyme complexes were formed by mixing CYTc-SpyCatcher and SpyTag-Enzymes, and the complexes retained their original enzymatic activity. Remarkably, the heme domain served as an electron acceptor from complexed enzymes by intramolecular electron transfer; consequently, all constructed CYTc-SC/ST-Enzyme complexes showed DET ability to the electrode, demonstrating the versatility of this method.

Wiring of bilirubin oxidases with redox polymers on gas diffusion electrodes for increased stability of self-powered biofuel cells-based glucose sensing.

A new redox polymer/bilirubin oxidase (BOD)-based gas diffusion electrode was designed to be implemented as the non-current and non-stability limiting biocathode in a glucose/O2 biofuel cell that acts as a self-powered glucose biosensor. For the proof-of-concept, a bioanode comprising the Os-complex modified redox polymer P(VI-co-AA)-[Os(bpy)2Cl]Cl and FAD-dependent glucose dehydrogenase to oxidize the analyte was used. In order to develop an optimal O2-reducing biocathode for the biofuel cell Mv-BOD as well as Bp-BOD and Mo-BOD have been tested in gas diffusion electrodes in direct electron transfer as well as in mediated electron transfer immobilized in the Os-complex modified redox polymer P(VI-co-AA)-[Os(diCl-bpy)2]Cl2. The resulting biofuel cell exhibits a glucose-dependent current and power output in the concentration region between 1 and 10 mM. To create a more realistic test environment, the performance and long-term stability of the biofuel cell-based self-powered glucose biosensor has been investigated in a flow-through cell design.

A New Digital Point-of-Care Tool With Advanced Blood Glucose Measuring Technology.

cobas® pulse is a point-of-care blood glucose (BG) measuring system for multiple-patient use in professional healthcare settings. The system provides advances in connectivity and BG measuring technology, and has multiple fail-safes to improve accuracy and reduce the risk of user error. Flavin adenine dinucleotide-dependent glucose dehydrogenase on the working electrode catalyzes oxidation of ß-D-glucose in the blood sample. A redox mediator/electron acceptor, on both the working and the counter electrode, facilitates diffusion of electrons in proportion to the glucose concentration and compensates for the effects of potential interfering agents. During development, >1 million test strip measurements were performed using >8000 test scenarios to refine the algorithm model. No clinically relevant interference was identified with extreme variations in blood properties and drugs in whole blood samples.